本文選題：黃芪多糖 + 乙型肝炎　； 參考：《四川農(nóng)業(yè)大學(xué)》2010年碩士論文

【摘要】： 乙型肝炎是由乙型肝炎病毒(hepatitis B virus, HBV)引起的一種影響人類(lèi)健康的世界性傳染病。接種乙肝疫苗是目前控制乙型肝炎感染的最有效的措施,但是傳統(tǒng)的乙型肝炎亞單位疫苗是由乙型肝炎病毒的表面抗原和鋁佐劑組成,鋁佐劑疫苗僅能引起很好的體液免疫反應(yīng)而不能引起有效的細(xì)胞免疫反應(yīng)。 佐劑可以增強(qiáng)細(xì)胞免疫和體液免疫,活化T細(xì)胞,上調(diào)細(xì)胞因子的表達(dá),增強(qiáng)免疫記憶,并能改變免疫反應(yīng)類(lèi)型。因此改良佐劑可能增強(qiáng)傳統(tǒng)乙型肝炎亞單位疫苗的免疫反應(yīng)。黃芪多糖(Astragalus polysaccharides,APS)是中藥黃芪的提取物,具有廣泛的藥理作用,也具有很強(qiáng)的免疫調(diào)節(jié)作用。APS可從多個(gè)方面發(fā)揮免疫調(diào)節(jié)作用：直接影響免疫細(xì)胞內(nèi)的物質(zhì)代謝,誘導(dǎo)產(chǎn)生相關(guān)的免疫調(diào)節(jié)因子；不僅能夠增強(qiáng)機(jī)體的特異性免疫反應(yīng),還可以增強(qiáng)非特異性免疫反應(yīng)；既可增強(qiáng)正常機(jī)體的免疫功能,又可調(diào)節(jié)異常機(jī)體的免疫功能。 本研究將APS作為乙型肝炎亞單位疫苗的佐劑與疫苗一起免疫昆明白小鼠,研究其對(duì)乙肝疫苗免疫反應(yīng)的影響,尤其是對(duì)細(xì)胞免疫反應(yīng)的影響。實(shí)驗(yàn)選取健康雌性6-8周齡昆明白小鼠35只,體重20-24 g,隨機(jī)分為5組,每組7只,分別為rHBsAg組、rHBsAg+APS組、rHBsAg+aulm組、APS組和對(duì)照組(不進(jìn)行免疫)。初次免疫設(shè)為0天,第14天加強(qiáng)免疫一次,在第21天進(jìn)行免疫學(xué)指標(biāo)的檢測(cè)。主要結(jié)果如下： 實(shí)驗(yàn)使用定量ELISA法檢測(cè)IgG水平,用MTT法檢測(cè)T細(xì)胞增殖反應(yīng),用流式細(xì)胞儀檢測(cè)體內(nèi)CTL反應(yīng),結(jié)果發(fā)現(xiàn)APS顯著增強(qiáng)乙肝疫苗誘導(dǎo)產(chǎn)生較高的IgG抗體水平,較強(qiáng)的T淋巴細(xì)胞增殖反應(yīng)和體內(nèi)CTL殺傷反應(yīng),表明APS能夠增強(qiáng)乙肝疫苗的免疫反應(yīng)。 為進(jìn)一步研究APS作為乙型肝炎亞單位疫苗的佐劑對(duì)免疫反應(yīng)的影響,對(duì)有關(guān)免疫學(xué)指標(biāo)作了進(jìn)一步檢測(cè)。用RT-PCR法檢測(cè)PFP、GraB、Fas L和Fas的表達(dá)以及用流式細(xì)胞儀檢測(cè)體內(nèi)細(xì)胞因子的表達(dá)水平,結(jié)果表明APS能夠增強(qiáng)乙肝疫苗上調(diào)PFP、GraB、Fas L、Fas和CD8+T細(xì)胞IFN-γ的表達(dá)水平,與體內(nèi)CTL反應(yīng)結(jié)果一致；APS能夠增強(qiáng)乙肝疫苗誘導(dǎo)較高的CD4+T細(xì)胞IFN-γ、IL-2和IL-4的表達(dá)水平,表明APS作為乙肝疫苗的佐劑能夠促進(jìn)機(jī)體Thl型和Th2型免疫反應(yīng)。 為進(jìn)一步探討APS作為佐劑增強(qiáng)免疫反應(yīng)的作用機(jī)理,用半定量PT-PCR法對(duì)TLR2、TLR4、TGF-β1和Foxp3的表達(dá)水平進(jìn)行檢測(cè),結(jié)果表明APS能夠很好地增強(qiáng)乙肝疫苗誘導(dǎo)TLR2和TLR4表達(dá),通過(guò)上調(diào)TLR2和TLR4的表達(dá)激發(fā)協(xié)同刺激分子和相關(guān)細(xì)胞因子的表達(dá)參與適應(yīng)性免疫應(yīng)答；APS作為乙肝疫苗的佐劑能夠明顯降低機(jī)體TGF-β1和Foxp3表達(dá)水平,通過(guò)抑制TGF-β1和Foxp3的表達(dá)促進(jìn)機(jī)體免疫反應(yīng)。 以上結(jié)果表明,APS作為乙型肝炎亞單位疫苗的佐劑能增強(qiáng)乙肝疫苗的免疫反應(yīng)。本研究初步證實(shí)了APS是一種良好的免疫佐劑,能夠顯著地提高乙型肝炎亞單位疫苗的體液免疫水平和細(xì)胞免疫水平,為乙型肝炎疫苗的研發(fā)提供了有意義的基礎(chǔ)研究數(shù)據(jù)資料。
[Abstract]:Hepatitis B is a worldwide infectious disease affecting human health caused by hepatitis B virus (HBV). Vaccination of hepatitis B is the most effective measure to control hepatitis B infection. However, the traditional hepatitis B subunit vaccine is composed of the surface antigen of hepatitis B virus and aluminum adjuvant, aluminum adjuvant. The vaccine can only cause good humoral immune response without inducing effective cellular immune response.
The adjuvant can enhance cell immunity and humoral immunity, activate T cells, increase the expression of cytokines, enhance the immune memory, and change the type of immune response. Therefore, the modified adjuvant may enhance the immune response of the traditional hepatitis B subunit vaccine. The Astragalus Polysaccharide (Astragalus polysaccharides, APS) is the extract of Astragalus membranaceus. Extensive pharmacological action and strong immunoregulation effect.APS can play an immunomodulatory role in many aspects: directly affecting the metabolism of substances in the immune cells and inducing related immunoregulatory factors; not only can enhance the specific immune response of the body, but also enhance the non specific immune response; it can enhance the normal function. The immune function of the body can also regulate the immune function of the abnormal body.
In this study, we immunized Kunming white mice with the adjuvant of APS as a subunit vaccine of hepatitis B, and studied the effect of the vaccine on the immune response of the hepatitis B vaccine, especially on the immune response of the hepatitis B vaccine. The experiment selected 35 healthy female 6-8 weeks old mice and weighed 20-24 G, randomly divided into 5 groups, each group of 7, respectively, group rHBsAg, rHBs Group Ag+APS, group rHBsAg+aulm, group APS and control group (no immunization). The first immunization was set for 0 days, the immunization was strengthened on the fourteenth day, and the immunological index was tested on the twenty-first day. The main results were as follows:
The experiment used quantitative ELISA to detect the IgG level, detect the proliferation reaction of T cells by MTT, and detect the CTL reaction in vivo by flow cytometry. The results showed that APS significantly enhanced the level of IgG antibody induced by hepatitis B vaccine, strong T lymphocyte proliferation reaction and in vivo CTL killing reaction, indicating that APS can enhance the immune reaction of hepatitis B vaccine. It should.
In order to further study the effect of APS as a adjuvant on hepatitis B subunit vaccine, the immune response was further examined. The expression of PFP, GraB, Fas L and Fas and the expression of cytokines in the body were detected by RT-PCR and the expression of cytokines in the body was detected by flow cytometry. The results showed that APS enhanced the hepatitis B vaccine to increase PFP and GraB, The expression level of IFN- gamma in Fas L, Fas and CD8+T cells is consistent with the results of CTL reaction in the body. APS can enhance the expression level of IFN- gamma, IL-2 and IL-4 of the hepatitis B vaccine, which indicates that the adjuvant of APS as a hepatitis B vaccine can promote the body type and the immune response of CD4+T.
In order to further explore the mechanism of APS as an adjuvant to enhance immune response, the expression level of TLR2, TLR4, TGF- beta 1 and Foxp3 was detected by semi quantitative PT-PCR method. The results showed that APS could enhance the expression of TLR2 and TLR4 induced by hepatitis B vaccine, and up up the table of TLR2 and TLR4 up to stimulate the table of synergistic stimulants and related cytokines. As an adjuvant of hepatitis B vaccine, APS can significantly reduce the expression level of TGF- beta 1 and Foxp3, and promote the immune response by inhibiting the expression of TGF- beta 1 and Foxp3.
The above results show that APS as a adjuvant of hepatitis B subunit vaccine can enhance the immune response of hepatitis B vaccine. This study preliminarily confirmed that APS is a good immune adjuvant. It can significantly improve the level of humoral immunity and cellular immunity of HBV subunit vaccine, which provides significance for the research and development of hepatitis B vaccine. Basic research data.

【學(xué)位授予單位】：四川農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2010
【分類(lèi)號(hào)】：R392

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本文編號(hào)：1895914